Amidation of Bioactive Peptides: The Structure of the Lyase Domain of the Amidating Enzyme

Eduardo E. Chufán; Mithu De; Betty A. Eipper; Richard E. Mains; L. Mario Amzel

doi:10.1016/j.str.2009.05.008

Amidation of Bioactive Peptides: The Structure of the Lyase Domain of the Amidating Enzyme

Eduardo E. Chufán, Mithu De, Betty A. Eipper, Richard E. Mains, L. Mario Amzel

Research output: Contribution to journal › Article › peer-review

42 Scopus citations

Abstract

Many neuropeptides and peptide hormones require amidation of their carboxy terminal for full biological activity. The enzyme peptidyl-α-hydroxyglycine α-amidating lyase (PAL; EC 4.3.2.5) catalyzes the second and last step of this reaction, N-dealkylation of the peptidyl-α-hydroxyglycine to generate the α-amidated peptide and glyoxylate. Here we report the X-ray crystal structure of the PAL catalytic core (PALcc) alone and in complex with the nonpeptidic substrate α-hydroxyhippuric acid. The structures show that PAL folds as a six-bladed β-propeller. The active site is formed by a Zn(II) ion coordinated by three histidine residues; the substrate binds to this site with its α-hydroxyl group coordinated to the Zn(II) ion. The structures also reveal a tyrosine residue (Tyr⁶⁵⁴) at the active site as the catalytic base for hydroxyl deprotonation, an unusual role for tyrosine. A reaction mechanism is proposed based on this structural data and validated by biochemical analysis of site-directed PALcc mutants.

Original language	English (US)
Pages (from-to)	965-973
Number of pages	9
Journal	Structure
Volume	17
Issue number	7
DOIs	https://doi.org/10.1016/j.str.2009.05.008
State	Published - Jul 15 2009
Externally published	Yes

Keywords

PROTEINS

ASJC Scopus subject areas

Structural Biology
Molecular Biology

Access to Document

10.1016/j.str.2009.05.008

Cite this

@article{c811ab9847fd429082235a9184c9b1a1,

title = "Amidation of Bioactive Peptides: The Structure of the Lyase Domain of the Amidating Enzyme",

abstract = "Many neuropeptides and peptide hormones require amidation of their carboxy terminal for full biological activity. The enzyme peptidyl-α-hydroxyglycine α-amidating lyase (PAL; EC 4.3.2.5) catalyzes the second and last step of this reaction, N-dealkylation of the peptidyl-α-hydroxyglycine to generate the α-amidated peptide and glyoxylate. Here we report the X-ray crystal structure of the PAL catalytic core (PALcc) alone and in complex with the nonpeptidic substrate α-hydroxyhippuric acid. The structures show that PAL folds as a six-bladed β-propeller. The active site is formed by a Zn(II) ion coordinated by three histidine residues; the substrate binds to this site with its α-hydroxyl group coordinated to the Zn(II) ion. The structures also reveal a tyrosine residue (Tyr654) at the active site as the catalytic base for hydroxyl deprotonation, an unusual role for tyrosine. A reaction mechanism is proposed based on this structural data and validated by biochemical analysis of site-directed PALcc mutants.",

keywords = "PROTEINS",

author = "Chuf{\'a}n, {Eduardo E.} and Mithu De and Eipper, {Betty A.} and Mains, {Richard E.} and Amzel, {L. Mario}",

note = "Funding Information: Mario Bianchet, Sandra Gabelli, and the staff at beamlines X4A and X6A of the National Synchrotron Light Source, Brookhaven National Laboratory, and of Advance Photon Source at Argonne National Laboratory are gratefully acknowledged for assistance during synchrotron data collection. Use of the Advanced Photon Source at Argonne National Laboratory was supported by the U.S. Department of Energy, Office of Science, Office of Basic Energy Sciences, under Contract No. DE-AC02-06CH11357. Use of the SGX Collaborative Access Team beamline facilities at Sector 31 of the Advanced Photon Source was provided by Eli Lilly & Company, who operates the facility. This work was supported by National Science Foundation grant MCB-0450465 (L.M.A.) and National Institutes of Health grant DK32949 (B.A.E. and R.E.M.). ",

year = "2009",

month = jul,

day = "15",

doi = "10.1016/j.str.2009.05.008",

language = "English (US)",

volume = "17",

pages = "965--973",

journal = "Structure",

issn = "0969-2126",

publisher = "Cell Press",

number = "7",

}

TY - JOUR

T1 - Amidation of Bioactive Peptides

T2 - The Structure of the Lyase Domain of the Amidating Enzyme

AU - Chufán, Eduardo E.

AU - De, Mithu

AU - Eipper, Betty A.

AU - Mains, Richard E.

AU - Amzel, L. Mario

N1 - Funding Information: Mario Bianchet, Sandra Gabelli, and the staff at beamlines X4A and X6A of the National Synchrotron Light Source, Brookhaven National Laboratory, and of Advance Photon Source at Argonne National Laboratory are gratefully acknowledged for assistance during synchrotron data collection. Use of the Advanced Photon Source at Argonne National Laboratory was supported by the U.S. Department of Energy, Office of Science, Office of Basic Energy Sciences, under Contract No. DE-AC02-06CH11357. Use of the SGX Collaborative Access Team beamline facilities at Sector 31 of the Advanced Photon Source was provided by Eli Lilly & Company, who operates the facility. This work was supported by National Science Foundation grant MCB-0450465 (L.M.A.) and National Institutes of Health grant DK32949 (B.A.E. and R.E.M.).

PY - 2009/7/15

Y1 - 2009/7/15

N2 - Many neuropeptides and peptide hormones require amidation of their carboxy terminal for full biological activity. The enzyme peptidyl-α-hydroxyglycine α-amidating lyase (PAL; EC 4.3.2.5) catalyzes the second and last step of this reaction, N-dealkylation of the peptidyl-α-hydroxyglycine to generate the α-amidated peptide and glyoxylate. Here we report the X-ray crystal structure of the PAL catalytic core (PALcc) alone and in complex with the nonpeptidic substrate α-hydroxyhippuric acid. The structures show that PAL folds as a six-bladed β-propeller. The active site is formed by a Zn(II) ion coordinated by three histidine residues; the substrate binds to this site with its α-hydroxyl group coordinated to the Zn(II) ion. The structures also reveal a tyrosine residue (Tyr654) at the active site as the catalytic base for hydroxyl deprotonation, an unusual role for tyrosine. A reaction mechanism is proposed based on this structural data and validated by biochemical analysis of site-directed PALcc mutants.

AB - Many neuropeptides and peptide hormones require amidation of their carboxy terminal for full biological activity. The enzyme peptidyl-α-hydroxyglycine α-amidating lyase (PAL; EC 4.3.2.5) catalyzes the second and last step of this reaction, N-dealkylation of the peptidyl-α-hydroxyglycine to generate the α-amidated peptide and glyoxylate. Here we report the X-ray crystal structure of the PAL catalytic core (PALcc) alone and in complex with the nonpeptidic substrate α-hydroxyhippuric acid. The structures show that PAL folds as a six-bladed β-propeller. The active site is formed by a Zn(II) ion coordinated by three histidine residues; the substrate binds to this site with its α-hydroxyl group coordinated to the Zn(II) ion. The structures also reveal a tyrosine residue (Tyr654) at the active site as the catalytic base for hydroxyl deprotonation, an unusual role for tyrosine. A reaction mechanism is proposed based on this structural data and validated by biochemical analysis of site-directed PALcc mutants.

KW - PROTEINS

UR - http://www.scopus.com/inward/record.url?scp=67649964485&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=67649964485&partnerID=8YFLogxK

U2 - 10.1016/j.str.2009.05.008

DO - 10.1016/j.str.2009.05.008

M3 - Article

C2 - 19604476

AN - SCOPUS:67649964485

SN - 0969-2126

VL - 17

SP - 965

EP - 973

JO - Structure

JF - Structure

IS - 7

ER -

Amidation of Bioactive Peptides: The Structure of the Lyase Domain of the Amidating Enzyme

Abstract

Keywords

ASJC Scopus subject areas

Access to Document

Other files and links

Fingerprint

Cite this